Mechanistic and therapeutic perspectives of non-coding RNA-modulated apoptotic signaling in diabetic retinopathy

Diabetic retinopathy (DR), a significant and vision-endangering complication associated with diabetes mellitus, constitutes a substantial portion of acquired instances of preventable blindness. The progression of DR appears to prominently feature the loss of retinal cells, encompassing neural retinal cells, pericytes, and endothelial cells. Therefore, mitigating the apoptosis of retinal cells in DR could potentially enhance the therapeutic approach for managing the condition by suppressing retinal vascular leakage. Recent advancements have highlighted the crucial regulatory roles played by non-coding RNAs (ncRNAs) in diverse biological processes. Recent advancements have highlighted that non-coding RNAs (ncRNAs), including microRNAs (miRNAs), circular RNAs (circRNAs), and long non-coding RNAs (lncRNAs), act as central regulators in a wide array of biogenesis and biological functions, exerting control over gene expression associated with histogenesis and cellular differentiation within ocular tissues. Abnormal expression and activity of ncRNAs has been linked to the regulation of diverse cellular functions such as apoptosis, and proliferation. This implies a potential involvement of ncRNAs in the development of DR. Notably, ncRNAs and apoptosis exhibit reciprocal regulatory interactions, jointly influencing the destiny of retinal cells. Consequently, a thorough investigation into the complex relationship between apoptosis and ncRNAs is crucial for developing effective therapeutic and preventative strategies for DR. This review provides a fundamental comprehension of the apoptotic signaling pathways associated with DR. It then delves into the mutual relationship between apoptosis and ncRNAs in the context of DR pathogenesis. This study advances our understanding of the pathophysiology of DR and paves the way for the development of novel therapeutic strategies. Graphical Abstract


Introduction
Diabetes mellitus is a prevalent global health disorder, with an estimated incidence of up to 285 million cases (Dhankhar et al. 2023).This condition is characterized by consistently high blood sugar levels due to insufficient insulin production by the pancreas or decreased sensitivity of cells to insulin.Type 1 diabetes, also known as insulin-dependent diabetes, is linked to inadequate insulin secretion, while Type 2 diabetes, known as insulin-independent diabetes, is associated with reduced cell responsiveness to insulin (Ozougwu et al. 2013).Hyperglycemia leads to various microvascular abnormalities in different parts of the body, including the retina, which can result in conditions like retinal detachment, neovascularization, and macular edema.These changes in the retina indicate damage caused by high blood sugar levels and the progression of diabetic retinopathy, a condition that significantly contributes to the risk of vision loss but can be preventable with appropriate interventions (Nentwich and Ulbig 2015).
The retina is a specialized tissue that plays a crucial role in light perception and the creation of visual this context, Wan et al. explored the protective influence exerted by miR-200a during the progression of DR.They observed reduced levels of miR-200a and elevated LIM domain protein 1 (PDLIM1) levels in both in vivo and in vitro DR models.Administration of miR-200a effectively mitigated retinal permeability and the expression of inflammatory factors.Furthermore, by targeting PDLIM1, miR-200a was observed to alleviate apoptotic conditions, enhance cell viability, and markedly reduce cellular migration in high-glucose-treated HRMECs.So, miR-200a presents itself as a prospective therapeutic target for the modulation of PDLIM1 expression DR (Wan et al. 2021).Similarly, recent evidence has revealed a direct interaction between miR-29 and the lncRNA RNA VIM-AS1.Also, exposure to high glucose (HG) resulted in upregulated miR-29 expression and downregulated VIM-AS1 levels.Furthermore, examination of cellular apoptosis demonstrated that the increased expression of VIM-AS1 mitigated the pro-apoptotic impact of miR-29 overexpression on h1RPE7 cells under HG conditions.Therefore, VIM-AS1 functions as a potential competitor for miR-29 binding, thereby influencing apoptotic regulation within h1RPE7 cells and potentially contributing to the development of DR (Zeng et al. 2020a).. Thereby, in the current paper, we initially present a detailed explanation of the apoptosis signaling pathway within the framework of DR.Subsequently, we elucidate the influence of ncRNAs on cellular apoptosis within the DR framework.Finally, in the concluding section, we illustrate the potential therapeutic utility of targeting ncRNAs/apoptosis axis in the management of DR (Fig. 1) (Table 1).

PI3K/AKT/mTOR in Retinal cells apoptosis
The PI3K/AKT/mTOR signaling cascade constitutes a pivotal pathway engaged in fundamental biological processes, including angiogenesis, apoptosis, proliferation, growth, and cellular metabolism (Karar and Maity 2011;Yu and Cui 2016).The PI3K/AKT signaling pathway entails a series of events, primarily involving the binding of extracellular factors to receptors, subsequent receptor activation leading to the phosphorylation of PI3K, PI3K-mediated phosphorylation of AKT, and the subsequent activation of downstream effector molecules.Following activation, PI3K facilitates PIP2 phosphorylation at the 3-position of the inositol ring, resulting in the production of PIP3 (Weernink et al. 2004).Subsequently, PIP3 functions as a docking site for two protein kinases, specifically AKT, also known as PDK1 (phosphoinositide-dependent protein kinase 1), and protein kinase B (PKB), which are recruited to the cellular membrane through their pleckstrin homology interaction domains (PH domains) (Cho and Park 2008;Powis et al. 2023).Upon being recruited to the cellular membrane, AKT is phosphorylated at Ser473 by mTORC2 (mTOR complex 2), which induces a structural alteration in AKT and facilitates its subsequent phosphorylation at Thr308 by PDK1 (Moore et al. 2011;Pullen et al. 1998).Upon being activated, AKT experiences a series of phosphorylation events on its target proteins.These initial phosphorylations take place at the cellular membrane, enabling the subsequent detachment of AKT.Subsequently, AKT relocates to the cytoplasm and nucleus, where it proceeds to phosphorylate further target proteins.Target proteins phosphorylation culminates in the activation of growth, cell survival, and proliferation (Chen et al. 2022).PI3K/AKT signaling pathway a critical influence on apoptosis within retinal cells, thereby potentially contributing to the pathogenesis of DR..In this regard, Zeng et al. documented a concurrent decline in DJ-1 protein levels and a rise in the apoptotic activity of RRPs within the HG group.Their further investigation revealed that exposure to HG concentrations for a duration of two days led to noteworthy levels of apoptosis in RRPs.This cellular condition demonstrated increased levels of ROS, along with an increase in p-p53 and activation of caspase-3.Furthermore, there was evidence of mitochondrial impairment, coupled with reduced catalase (CAT) and manganese superoxide dismutase (MnSOD) activities.Furthermore, a decline in DJ-1 protein expression, along with diminished levels of its subsequent targets, phosphorylation state of AKT, and mTOR was evident.Conversely, elevation of DJ-1/PARK7 led to contrasting outcome.Additionally, it enhanced  activities of MnSOD and CAT, thus improving mitochondrial functionality.Furthermore, the increased expression of DJ-1/PARK7 was linked to a decrease in the expression of genes involved in apoptosis, specifically p-p53 and activated caspase-3.This led to a decline in ROS production and a lower rate of apoptosis in renal proximal tubular cells exposed to HG levels.These findings posit a protective role for DJ-1 in shielding retinal pericytes from oxidative stress damage induced by HG exposure.Consequently, DJ-1 has the potential to augment mitochondrial function, mitigate ROS generation, and bolster antioxidant capacity, thereby mitigating apoptosis in retinal pericytes.This effect might be facilitated via the PI3K/AKT/ mTOR signaling pathway, which possibility associated with the initial pathophysiological processes DR (Zeng et al. 2019).

TLR4/NF-κB in Retinal cells apoptosis
The TLR4/NF-κB pathway is a pivotal inflammatory signaling cascade intricately associated with cellular apoptosis, proliferation, differentiation, and the activation of pro-inflammatory responses.Tolllike receptors (TLRs) constitute a category of receptors that are classified within transmembrane proteins known as pattern recognition receptors (PRRs) (Kawai and Akira 2010;Zhu et al. 2018a).Among mammals, PRRs are unique molecules capable of transmitting extracellular antigenic signals to cells, thereby initiating an inflammatory response.TLR4 holds the distinction of being the inaugural member identified within the Toll-like receptor family, and it functions in both immune regulation and immune defense (Zhang et al. 2022).When activated, TLR4 undergoes dimerization and initiates two primary signaling pathways: one dependent on myeloid differential factor 88 (MyD88) and the other on toll/ interleukin 1 receptor domain-containing adaptor inducing interferon-beta (TRIF).These signaling cascades subsequently lead to the activation of downstream factors such as mitogen-activated protein kinases (MAPKs), and NF-κB, ultimately inducing a diverse array of pro-inflammatory genes, including cytokines and enzymes associated with inflammation (Yu et al. 2022;Fitzgerald et al. 2001;Piras and Selvarajoo 2014).Recent experimental findings suggests that the TLR4/NF-κB signaling pathway assumes a pivotal role in the regulation of apoptosis in retinal cells and contributes significantly to the progression of DR.In this context, according to recent experimental findings, HG elevated the expression levels of TLR4, and four downstream signaling molecules associated with TLR4 (NLRP3, TRAF6, NF-κB, and MyD88) and pro-inflammatory cytokines (IL-18 and IL-1β) in retinal ganglion cells (RGCs).Notably, HG trigger a conspicuous elevation in the apoptosis of RGCs.Furthermore, TAK-242 introduction, which functions as an antagonist of TLR4, effectively suppressed both inflammation and RGC apoptosis within the HG group.These findings unequivocally illustrated the pivotal involvement of TLR4 in the inflammatory response and apoptotic processes of RGCs triggered by HG (Hu et al. 2017) (MDA) content, and decreased NADPH oxidase 4 expression, while enhanced the activities of GPX, catalase, and superoxide dismutase under HG conditions.So, miR-455-5p significantly attenuated oxidative stress injury induced by HG.Most importantly, miR-455-5p directly target suppressor of cytokine signaling 3 (SOCS3), and miR-455-5p exerts a negative regulatory influence on the expression of SOCS3.Moreover, SOCS3 restoration nullified the advantageous impacts of miR-455-5p on apoptosis and the buildup of ROS.Taken together, miR-455-5p mitigated HG-induced damage by inhibiting apoptosis and oxidative stress through the targeting of SOCS3, suggesting that miR-455-5p emerges as a promising candidate for future DR therapeutic development (Chen et al. 2019).

MiR-29a/b
Within the human genome, the miR-29 family comprises four members: miR-29b-1, miR-29b-2, miR-29a, and miR-29c.These miR-NAs originate from two distinct primary transcripts: pri-miR-29a/b1 cluster located on chromosome 7q32.3and pri-miR-29b2/c cluster situated on chromosome 1q32.2(Horita et al. 2021).The miR-29 family is frequently associated with DM in various studies.In diabetic patients, miR-29a expression shows downregulation, while STAT3 and IL-6 levels exhibit upregulation in both the lens capsules and aqueous humor.As well, Overexpression of miR-29a combined with si-STAT3 transfection mitigated the increase in apoptosis and the disruption of MMP caused by HG.Notably, suppression of STAT3 resulted in an elevated Bcl-2/Bax ratio when compared to cells subjected to HG treatment alone.As well, it was observed that STAT3 upregulation countered the impacts of miR-29a transfection.Furthermore, by suppressing STAT3 signaling pathway, miR-29a alleviates the adverse impacts of HG on cell viability and mitochondrial dysfunction.Also, upregulation of miR-29a led to a partial decrease in ROS generation, elevated MDA levels, reduced SOD activity, and an upregulation of the Bcl-2/Bax ratio, which are linked to apoptotic pathways.These findings underscore miR-29a involvement in oxidative damage and apoptotic processes.In this manner, HG induces inflammatory responses and initiates mitochondrial dysfunction, thereby augmenting apoptosis via activation of the IL-6/STAT3 pathway (Li et al. 2021b).On the other hand, numerous studies have consistently shown that miR-29 functions as a proapoptotic factor in this particular condition.In this context, HG resulted in a concentration-dependent increase in apoptotic cell death within RPE cells, and miR-29 exhibited an upregulation in response to HG within RPE cells.Additionally, Downregulation of miR-29 expression mitigated HG-induced apoptosis in RPE cells, as evidenced by a reduction in caspase-7 protein generation.Also, miR-29 directly target PTEN, and downregulation of miR-29 increased PTEN expression.Thereby, miR-29 contributes to the apoptotic pathway triggered by HG in RPE cells.This effect is likely mediated by its inverse association with PTEN expression (Lin et al. 2016).Furthermore, Zeng et al. discovered that the miR-29b-3p expression exhibited a 3.2-fold increase, while the SIRT1 protein was reduced in patients with DR.In the HG-CoCl2 environment, they observed an elevation in miR-29b-3p and the Bax/Bcl-2 ratio, accompanied by a reduction in SIRT1 within HRMECs.They observed a decrease in the viability of HRMECs and a concomitant rise in apoptotic cell death when exposed to HG-CoCl2 treatment.Ultimately, they demonstrated that miR-29b directly target SIRT1, and that the Upregulation of miR-29b-3p corresponded with a significant down-regulation of SIRT1 protein expression and a concomitant increase in the Bax/Bcl-2 ratio.Conversely, down-regulation of miR-29b-3p resulted in opposite effects.Furthermore, SRT1720, a SIRT1 agonist, mitigated the apoptosis of HRMECs induced by miR-29b-3p by enhancing the expression of SIRT1 protein.So, aberrant regulation of miR-29b-3p/SIRT1 axis is a crucial mechanism driving the apoptosis of HRMECs in DR (Zeng et al. 2020).

MiR-204
The miR-204 resides within the sixth intron of the transient receptor potential melastatin 3 (TRPM3) gene.. MiR-204 assumes a pivotal role in both the developmental processes and functional aspects of the retina, with its dysregulation being implicated in the pathogenesis of various retinopathies (Bereimipour et al. 2021).Quantitative analysis of miR-204 expression in the retinal tissues of DR model rats revealed a significant downregulation in comparison to healthy controls.This observation suggests a dysregulation of miR-204 in DR pathogenesis, potentially contributing to the development of the disease.Moreover, Transfection with miR-204 mimics resulted in a marked upregulation of endogenous Page 11 of 32 53 Vol.: (0123456789) miR-204 expression.This, in turn, mediated a coordinated increase in the expression of Bcl-2 and SIRT1, while concurrently suppressing the production of proinflammatory mediators like TNF-α.Thus, miR-204 mitigates inflammation and cellular apoptosis in DR in rats via an increase in Bcl-2 and SIRT1 expression (Qi et al. 2020).

MiR-211
MiR-211 is produced through the transcriptional process of Trpm1 gene.It is important to note that Trpm1 gene is located on chromosome 15q13-q14, a genomic region commonly associated with deletions found in various cancerous conditions.Furthermore, MiR-211 is situated specifically within the sixth intron of the Trpm1 gene (Yuan et al. 2022;Ye et al. 2022).MiR-211 may play a pivotal role in regulating the metabolic and catabolic processes of retinal cells, with its importance extending to the maintenance of adult visual functionality (Barbato et al. 2017).Diabetic cataract (DC) mice exhibited elevated levels of p53, Bax, and miR-211, alongside reduced levels of SIRT1 and Bcl-2.As well, miR-211 functions as a direct regulator of SIRT1 expression.Furthermore, upregulation of miR-211 trigger elevated expressions of p53, Bax, and miR-211 alongside reduced expressions of SIRT1 and Bcl-2.Notably, elevated expression of miR-211 led to decreased proliferation, and increased apoptosis of lens epithelial cells, while miR-211 depletion is associated with opposite effects.Therefore, miR-211 may function as a regulator of lens epithelial cell homeostasis in DC mice.miR-211 overexpression appears to promote apoptosis and inhibit proliferation, potentially through its interaction with SIRT1.These findings implies a potential therapeutic role for miR-211 in the treatment of DR (Fitzgerald et al. 2001).Vol:.( 1234567890) MiR-365 Zheng et al. explored miR-365 influence on retinal neuron apoptosis in rats with diabetes mellitus, primarily through its targeting of IGF-1.In their research, suppression of miR-365 through the administration of anti-miR-365 led to a reduction in apoptotic cells and Bax protein levels relative to control groups.Conversely, the groups treated with sh-IGF-1 and anti-miR-365 + sh-IGF-1 exhibited a contrasting pattern of results.Furthermore, the diabetes mellitus rat models displayed elevated levels of miR-365 and Bax expression, alongside reduced levels of Bcl-2 and IGF-1 expression.Additionally, in the diabetic rat models, a higher quantity of cells undergoing apoptosis was noted.Furthermore, they demonstrated that the sh-IGF-1 group exhibited decreased Bax expression, increased Bcl-2 and IGF-1 expressions, and a reduced number of apoptotic cells.Lastly, they observed that the anti-miR-365 + sh-IGF-1 groups displayed an upregulation of Bax expression, downregulation of IGF-1 and Bcl-2 expressions, and an increased count of apoptotic cells when compared to the anti-miR-365 group.Their findings indicate that the inhibition of miR-365 results in heightened expression of IGF-1, which subsequently exerts anti-apoptotic effects on retinal neurons in diabetic rats.These results underscore the potential of miR-365 as a viable therapeutic target in the context of DR (Zheng et al. 2018).

MiR-146a
The miR-146a gene is situated on human chromosomes 19, 10, and 5, whereas in mice, it is located on chromosome 11.A substantial body of medical research has consistently identified dysregulation of miR-146a in individuals with T2D, underscoring the significant role of miR-146a in the development of T2D and its associated complications (Ghaffari et al. 2023) of miR-542-5p effectively curbed apoptosis in RPE cells subjected to HG conditions.Furthermore, miR-542-5p was found to directly interact with coactivator-associated arginine methyltransferase 1 (CARM1).Overexpression of miR-542-5p caused a decrease in CARM1 expression levels, whereas suppression of miR-542-5p led to an elevation in CARM1 levels.Importantly, upregulation of CARM1 promoted apoptosis mediated by miR-542-5p in RPE cells exposed to HG conditions.Thus, miR-542-5p's direct targeting of CARM1 suggests a protective role for miR-542-5p in RPE cells, opening new avenues for therapeutic interventions in DR (Guo et al. 2020).

MiR-133b
The miR-206/miR-133b cluster is remarkably preserved in the muscle tissues of various organisms, including flies, mice, and humans, and it is situated on chromosome 6p12.2(Nohata et al. 2012).Yao et al. noted that texposure to HG or miR-133b silencing result in a decrease in apoptosis in hRECs.Furthermore, HG resulted in an increase in the mRNA and protein levels of LIMK, ROCK1, RhoA, and the p-MLC protein in hRECs.On contrary, elevated expression of miR-133b suppressed cellular proliferation, enhanced apoptosis, and led to a downregulation of both mRNA and protein quantities of RhoA, ROCK1, LIMK, and p-MLC in hRECs subjected to HG conditions.Thereby, heightened expression of miR-133b hindered cell proliferation and stimulated apoptosis in a DR cell model through the reduction of RhoA expression (Yao et al. 2018).

MiR-499-3p
Liu et al. demonstrated a decrease in IFNA2 levels and an increase in miR-499-3p expression in retinal tissues and cells of diabetic rats.This molecular alteration resulted in the activation of the TLR4 signaling pathway.They ascertained that miR-499-3p function as a direct regulator of IFNA2, and exerting an inhibitory effect on its expression.Also, miR-499-3p may function as a potential regulator of the TLR4 signaling pathway by targeting IFNA2.
Their further experiments revealed that the reduction in miR-499-3p levels facilitated the proliferation of retinal cells and concurrently inhibited apoptosis, thereby mitigating the effects of DR.In conclusion, miR-499-3p facilitated the progression of retinopathy by activating TLR4 signaling pathway and promoting apoptosis, underscoring its potential for exploration as a therapeutic target for further research in DR (Liu et al. 2020).

MiR-34a
Residing on chromosome 1p36, miR-34a) has been implicated in the pathophysiology of DM (Mone et al. 2022).MiR-34a upregulated, while SIRT1 downregulated in retinal tissues of DR-afflicted rats and in HG-induced RVECs.Suppression of miR-34a had a beneficial impact on DR, exerting its influence by modulating apoptosis and the expression of VEGF in DR rats.As well, miR-34a directly target SIRT1, and overexpression of miR-34a suppressed proliferation and induced apoptosis in RVECs, and these effects successfully reversed by increasing SIRT1 expression.Therefore, miR-34a drives apoptosis in RVECs by modulating SIRT1 in DR rats, underscoring the potential significance of the miR-34a/SIRT1 axis as a potential avenue for DR treatment (Ji et al. 2020).Furthermore, Ma et al. noted that retinal cell apoptosis, the expression levels of miR-34α, NF-κB, HMGB1, and caspase-3 increased in rats with DR, while miR-34α silencing exert opposite effects..In this manner, suppression of miR-34α mitigates retinal cell apoptosis in rats with DR by modulating the expression of HMGB1 and the downstream NF-κB pathway (Ma et al. 2020).12% of all identified microRNA genes (Liu et al. 2019a).Zhang et al. explored the miR-203a-3p function in CoCl2-induced apoptosis of RPEs.They demonstrated the expression of miR-203a-3p within the RPE of both control and diabetic rat models, and identified miR-203a as a direct modulator of SOCS3.They disclosed that the introduction of miR-203a-3p mimics led to an enhancement in CoCl2-induced apoptosis of RPEs.However, the apoptotic effects of miR-203a-3p were partially counteracted by the increased expression of SOCS3 or the administration of the JNK inhibitor SP600125.So, miR-203a-3p functions as a significant novel regulator of apoptosis induced by CoCl2 in RPE cells, operating by regulating the expression of SOCS3.In conclusion, deregulation of miR-203a-3p/SOCS3/JNK/c-Jun signaling pathway is hypothesized to be a critical contributor to RPE cell apoptosis in DR (Zhang et al. 2020).

MiR-296-5p
MiR-296 is situated within the chromosomal region 20q13.32,exhibiting substantial sequence conservation across species and exerting significant roles in various biological processes, including apoptosis (Zhu et al. 2018b).In DR mice, retinal tissues exhibited a marked increase in G protein subunit alpha i2 (GNAI2) and HDAC3 expression levels, accompanied by a significant decrease in miR-296-5p expression.The researchers noted that restoration of miR-296-5p or HDAC3 silencing resulted in a decrease in Evans blue leakage in DR mice.Additionally, it mitigated retinal ganglion cells apoptosis, decreased levels of MDA and VEGF, and increased SOD activity in both retinal tissues and serum of DR mice.Through interaction with the miR-296-5p promoter region, HDAC3 functioned as a suppressor of miR-296-5p expression, leading to a subsequent upregulation of GNAI2.In this manner, reducing HDAC3 levels or reinstating miR-296-5p mitigates the apoptosis of RGCs in DR mice by lowering the expression of GNAI2 (Che et al. 2022).

MiR-345-5p
MiR-345, a small nvRNA, is situated on human chromosome 14q32.2and has been implicated in diverse human pathological conditions (Natesh et al. 2021) This novel insight contributes to our understanding of the initiation and progression of DR (Luo et al. 2020).Additionally, Tong et al. investigated the signaling pathways underlying MEG3-mediated regulation of inflammation and apoptosis in DR.Within their study, HG exposure exhibited suppressive effects on SIRT1 and MEG3 expression, while concurrently inducing an upregulation of miR-34a levels.Their investigation unveiled that MEG3 could enhance the expression of SIRT1 through its interaction with miR-34a.Furthermore, HG-induced apoptosis and the release of inflammatory cytokines, including TNFα, IL-6, and IL-1β, were attenuated by the upregulation of MEG3 and miR-34a silencing.However, the miR-34a upregulation counteracted these protective effects mediated by MEG3.So, overexpression of MEG3 resulted in the inhibition of the NF-κB signaling pathway and an elevation in the Bcl-2/Bax ratio, which was achieved through the downregulation of miR-34a.Thereby, MEG3 demonstrated the capability to mitigate apoptosis and inflammation induced by HG by suppressing the NF-κB signaling pathway via its interaction with miR-34a/SIRT1 axis.Therefore, MEG3/miR-34a/SIRT1 axis represents a promising target for therapeutic intervention in DR (Tong et al. 2019).
LncRNA PVT1/miRNA axis Experimental finding from Yang et al. unveiled an elevation in the expression of lncRNA PVT1 within the HLE B-3 cells induced by HG in contrast to the control group maintained under normal glucose conditions.Furthermore, they provided evidence indicating that the transcription factor SP1 exhibit potential DNA binding activity to PVT1 promoter region, thereby facilitating its transcription.Their functional experiments revealed that the suppression of PVT1 had the capacity to hinder the proliferation and enhance the apoptotic processes in HLE B-3 cells.In their mechanistic inquiry, they disclosed that PVT1 operated as a 'miRNA sponge,' directing its regulatory effects towards the miR-214-3p/MMP2 axis.Their discovery has unveiled a fresh perspective on the involvement of the lncRNA PVT1/miR-214-3p/MMP2 axis in the pathogenesis of DC, offering valuable insights into the mechanisms underlying this condition (Yang et al. 2020).In addition, Guo et al. demonstrated Page 17 of 32 53 Vol.: (0123456789) that PVT1 exhibited increased expression in cellular models of DR, and the inhibition of PVT1 resulted in enhanced cell viability and reduced apoptosis in HGtreated ARPE-19 cells.Their findings additionally demonstrated that PVT1 could directly interact with miR-1301-3p, exerting a negative regulatory effect on miR-1301-3p expression.Subsequent experiments unveiled that miR-1301-3p directly targeted KLF7, and PVT1 elevated KLF7 expression through its interaction with miR-1301-3p.They finally confirmed that the impact on cell viability and apoptosis resulting from PVT1 silencing could be restored through KLF7 overexpression.Therefore, PVT1demonstrates a suppressive impact on the proliferation and induces apoptosis in HG-treated ARPE-19 cells by interacting with miR-1301-3p, which subsequently leads to the upregulation of KLF7 (Guo et al. 2022).LncRNA XIST/miRNA axis Wang et al. substantiated the interaction between miR-34a and both XIST and SMAD2 using a luciferase reporter assay.Their study revealed an upregulation in XIST expression and a concurrent downregulation in miR-34a levels within DC tissues and SRA01/04 cells subjected to HG treatment.They additionally showed that suppressing XIST expression or enhancing miR-34a levels resulted in reduced cell proliferation, while inducing apoptosis in SRA01/04 cells exposed to HG conditions.It was determined that miR-34a directly interacted with SMAD2, and XIST positively regulated the expression of SMAD2.Further, in SRA01/04 cells exposed to HG, XIST silencing suppressed proliferation and enhanced apoptosis, which were however mitigated by SMAD2 upregulation.

LncRNA MIAT/miRNA axis
In conclusion, XIST was identified as a promoter of cell proliferation and an inhibitor of apoptosis in DC, functioning through the regulatory axis of miR-34a/ SMAD2.These findings propose the XIST/miR-34a/SMAD2 axis could serve as a potential novel biomarker for the management of DC (Wang et al. 2022a).AK077216 specifically in DR patients, as opposed to diabetic patients without evident complications, when compared to healthy controls.In their study, the reduced expression of AK077216 served as a distinguishing feature that set apart DR patients from both diabetic patients without complications and healthy controls.Furthermore, treatment with HG did not significantly affect lncRNA AK077216 expression in ARPE-19 cells.Their investigation revealed a reciprocal relationship between miR-383 and lncRNA AK077216 in DR patients, where the upregulation of lncRNA AK077216 led to a decrease in miR-383 levels.However, the upregulation of miR-383 did not induce a significant alteration in the expression of lncRNA AK077216.Additionally, their research revealed that the upregulation of lncRNA AK077216 had an inhibitory effect on apoptosis in ARPE-19 cells, whereas the upregulation of miR-383 exerted an opposing role, mitigating the impact of lncRNA AK077216 upregulation.Therefore, LncRNA AK077216 exhibits downregulation in the context of DR and exerts an inhibitory effect on apoptosis in ARPE-19 cells by means of the downregulation of miR-383 (Zhang et al. 2019).

LncRNA AK077216/miRNA axis
LncRNA SNHG16/miRNA axis Li et al. reported elevated expression of E2F1 and SNHG16, alongside reduced miR-20a-5p levels, in proliferative DR compared to control or non-proliferative DR conditions.They discovered that SNHG16 functions as a regulatory hub, modulating miR-20a expression and its interactions with both SNHG16 and E2F1.Subsequent experiments demonstrated that overexpression of SNHG16 via plasmid transfection promoted both cell apoptosis and vessel-like formation, whereas introduction of a miR-20a-5p mimic partially abrogated these effects.They ultimately showed that the introduction of a gene silencing E2F1 plasmid effectively reversed the exacerbating effects of SNHG16 overexpression on proliferative DR.In conclusion, SNHG16 as a miR-20a-5p sponge, demonstrating its regulatory control over E2F1 expression and a consequent exacerbation of apoptosis in proliferative DR (Li et al. 2021c).
LncRNA RPSAP52/miRNA axis Niu et al. explored the roles of RPSAP52 in DR.Their findings indicated a notable reduction of RPSAP52 in DR patients in comparison to individuals with diabetes lacking evident complications.They revealed a direct interaction between RPSAP52 and miR-365, but the upregulation of miR-365 and RPSAP52 did not lead to any significant alteration in each other's expression.They also demonstrated that the RPSAP52 overexpression resulted in the upregulation of TIMP metallopeptidase inhibitor 3 (TIMP3) in RPE cells.HG treatment of RPE cells led to decreased expression of TIMP3 and RPSAP52, while miR-365 levels were conversely upregulated.Moreover, through cell apoptosis analysis, exposure to HG treatment in RPE cells resulted in a significant upregulation of both TIMP3 and RPSAP52, which was associated with a decreased rate of apoptosis.Therefore, RPSAP52 acts as an endogenous sponge for miR-365, promoting TIMP3 expression and consequently inhibiting RPE cell apoptosis in DR (Niu et al. 2020).
LncRNA FLG-AS1/miRNA axis Compared to healthy controls, DR patients exhibited a significant decrease in FLG-AS1 expression and a notable increase in miR-380-3p expression in their serum.
Further, the expression levels of miR-380-3p, and FLG-AS1 exhibited a negative correlation among DR patients.Moreover, upregulating FLG-AS1 protected ARPE-19 cells from HG-induced apoptosis, and mitigated retinal injury in diabetic rats.Mechanistically, FLG-AS1 directly targets miR-380-3p, thereby promoting SOCS6 expression.Importantly, either suppression of miR-380-3p or upregulation of SOCS6 mitigated apoptosis in ARPE-19 cells exposed to HG conditions Thereby, FLG-AS1 acts as a protective factor in DR by regulating the miR-380-3p/SOCS6 axia, thereby mitigating apoptosis in retinal epithelial cells (Luo et al. 2022).finally substantiated that SNHG4 and Oxr1 exerted contrasting effects and counteracted the outcomes associated with the upregulation of miR-200b.In conclusion, SNHG4 potentially acts as a sponge for miR-200b, thereby restraining cell apoptosis in DR through the upregulation of Oxr1 (Yu et al. 2021).

LncRNA SNHG4/miRNA axis
LncRNA MALAT1/miRNA axis Li et al. explored the regulatory mechanisms involving MALAT1, miR-378a-3p, and PDE6g in apoptosis among RMECs exposed to HG conditions.HG induced a concomitant upregulation of PDE6G and MALAT1, while conversely inhibiting miR-378a-3p expression.Subsequent experiments revealed that MALAT1 overexpression facilitated the proliferation of RMECs and concurrently suppressed apoptosis when exposed to HG conditions.In their study, MALAT1 competitively sequestered miR-378a-3p, a molecule that targeted PDE6G.Consequently, the MALAT1/miR-378a-3p/PDE6G signaling axis functions to mitigate apoptosis in RMECs under HG conditions (Li LncRNA MCM3AP-AS1/miRNA axis Recent investigation disclosed that MCM3AP-AS1 reduced in DR patients in comparison to individuals with type 2 diabetes mellitus who did not exhibit significant complications.As well, exposure to HG led to the downregulation of MCM3AP-AS1 in RPEs.Notably, according to cell apoptosis analysis overexpression of MCM3AP-AS1 and SIRT1 resulted in a decrease in the apoptotic rate of RPEs.However, the upregulation of miR-211 partially counteracted the effects of MCM3AP-AS1 and SIRT1 overexpression.In this manner, MCM3AP-AS1 exhibits reduction in DR and facilitates cell apoptosis through the regulation of the miR-211/SIRT1 pathway (Xia et al. 2022a) (Fig. 4).

CircRNA/miRNA/mRNA axis-mediated apoptosis in DR
CircRNAs, recognized as emerging molecules of significant interest, are found in high abundance within various eukaryotic cells.They are transcribed from a substantial number of genes in both human and animals (Zhou et al. 2018).The majority of circRNAs originate from the backsplicing of exons or introns derived from pre-messenger RNA (pre-mRNA) genes responsible for encoding proteins.This process leads to the connection of the downstream 5' splicing site with the upstream 3' splicing site, resulting in the formation of a circular RNA molecule held together by a 3'-5'-phosphodiester bond (Welden and Stamm 2019).It is widely recognized that a subset of circR-NAs, which contains MREs, can function as competitive molecules.These circRNAs interact with various miRNAs, thereby influencing the efficacy of miRNAs in regulating downstream mRNA expression (Su et al. 2024).Multiple investigations have provided evidence of circRNA-miRNA-mRNA regulatory pathways contributing to the apoptosis of retinal cells and the development of DR.Thereby, in the following section we will explore the current state of knowledge regarding the regulatory interactions between cir-cRNA, miRNA, and mRNA in retinal cells apoptosis and DR.
CircZNF532/miRNA axis Liang et al. demonstrated that DR patients and and HG-induced ARPE-19 cells displayed upregulated STAT3 and circ-ZNF532 expression, alongside decreased miR-20b-5p levels.They observed that increased miR-20b-5p levels or circZNF532 silencing resulted in heightened proliferation and diminished apoptosis in ARPE-19 cells.They further established that circZNF532 function as a ceRNA for miR-20b-5p, subsequently suppressing its expression.Additionally, miR-20b-5p was validated as a direct regulator of STAT3.Their findings demonstrated that miR-20b-5p directly targets STAT3, thereby regulating the viability and apoptosis of ARPE-19 cells.Their final analysis revealed elevated levels of STAT3 and circZNF532 in mice with STZ-induced diabetes, accompanied by a significant decrease in miR-20b-5p expression relative to the control group.Notably, they noted that circZNF532 silencing resulted in the suppression of apoptosis in the retinal tissues of mice.Thereby, circZNF532 overexpression may exacerbate the development of DR by undermining the suppressive action of miR-20b-5p on STAT3, offering novel insights into the pathogenesis of DR and advocating for further exploration of the circZNF532/miR-20b-5p/STAT3 axis as a promising candidate for therapeutic intervention in this condition (Liang et al. 2022). in the context of DC.In their study, the levels of circ-PAG1 were found to be diminished in individuals withDC, and the increased expression of circPAG1 mitigated the inhibitory effects of HG on cell viability and proliferation, while also counteracting the promotion of cell apoptosis and oxidative stress.They noted that circPAG1 acted as a miR-211-5p sponge, and the protective function of circPAG1 was partially accomplished through its sequestration of miR-211-5p.They also revealed that miR-211-5p targeted E2F3, and circPAG1 increased the expression of E2F3 by sequestering miR-211-5p.In their final analysis, they provided evidence that miR-211-5p suppression alleviated HG-induced cell apoptosis and oxidative stress by elevating E2F3 expression.In this manner, their study elucidated that circPAG1 conferred protection to human lens epithelial cells against HG-induced cell apoptosis and oxidative stress through its regulation of the miR-211-5p/E2F3 axis (Tao et al. 2022).(Natesh et al. 2021) (Fig. 5).

Targeting apoptosis in DR: from Herbal medicine to novel therapy
Emerging evidence underscores the significant contribution of apoptosis in retinal cells, particularly retinal ganglion cells, to the development of DR.This suggests the potential for apoptosis as an alternative therapeutic strategy to address DR.At present, there exist various compounds primarily focused on modulating ncRNAs to inhibit apoptosis in DN.The following section provides an overview of these strategies and agents.
Cell Biol Toxicol

Herbal medicine
Herbal medicines are categorized as products derived from plants and employed for the purpose of maintaining or restoring health, as defined by the National Institutes of Health.Historical records indicate that herbal medicine has been employed for more than 5000 years, serving as the sole documented form of medicine during that era (Gavanji et al. 2023).Contemporary scholarly literature presents compelling evidence regarding the potential efficacy of herbal medicine as agents against DR.The active constituents within herbal medicines can interact with a variety of ncRNAs, including miRNAs, lncRNAs, and circRNAs, which are energing as promising therapeutic targets for a wide range of diseases.Thereby, these molecules, acting through ncRNAs, can modulate regulatory processes, particularly those related to pro-apoptotic pathways, and thereby offer protective effects in various pathological contexts, including diabetes.In the following section, we focus on herbal extracts that target ncRNAs and apoptosis in DR.
NcRNA-mediated apoptosis modulation via Astragaloside-IV in DR Astragaloside IV (AS-IV) is a naturally occurring saponin derived from Radix astragali, commonly known as Huangqi in traditional Chinese medicine.This compound has gained prominence for its growing recognition of possessing potential anti-apoptotic, antioxidant, and antiinflammatory, attributes.Over the past decade, an accumulating body of evidence has underscored the protective effects of AS-IV against DM (Li et al. 2023).In a study by Wang et al They finally found that beneficial effects of APS on RPE cells subjected to HG conditions were negated by miR-195 upregulation.In this manner, APS effectively mitigated the metabolic memory phenomenon in RPE cells exposed to HG by preventing mitochondrial dysfunction-induced apoptosis through the regulation of miR-195 (Liu et al. 2019b).Moreover, Gao et al. utilized ARPE-19 cell lines and primary RPE cells to empirically assess the impact of APS on mitochondrial damage and apoptosis triggered by metabolic memory resulting from HG exposure.They observed that a regimen of HG followed by NG exposure significantly elevated the levels of miR-182 and concurrently reduced the expression of its target gene, Bcl-2.APS treatment effectively counteracted these aforementioned effects.They additionally noted that APS treatment reinstated proper mitochondrial function and mitigated apoptosis in cells exhibiting metabolic memory.In their final observation, they found that the protective effects of APS against mitochondrial and apoptosis were partially diminished upon upregulation of miR-182.In this manner, APS mitigated mitochondrial damage and apoptosis induced by metabolic memory through the regulation of the miR-182/Bcl-2 axis, potentially offering a novel therapeutic approach for managing DR (Gao et al. 2021).
NcRNA-mediated apoptosis modulation via blueberry anthocyanin extract in DR Blueberries are recognized as a nutritious fruit due to their high anthocyanin content.An increasing body of research indicates that anthocyanins derived from the berries of blueberry plants offer significant nutritional and health benefits.Beyond their widely recognized antioxidant properties, the prominent active constituents in blueberries, specifically anthocyanins, exhibit notable anticancer and anti-type 2 diabetes properties (Wu et al. 2023)

Novel therapy in DR
For a long time, it has been documented that cells release vesicles into the extracellular space.These extracellular vesicles (EVs) come in various forms, which are distinguished by their size and the mechanisms underlying their formation.There exist three recognized categories of EVs: microvesicles, apoptotic bodies, and exosomes.Among these, exosomes are released by a wide range of cell types.Exosomes are involved in both normal and pathological cellular interactions by facilitating the transfer of RNA, lipids, and proteins.The composition and impact of exosomes are contingent on the characteristics of the donor cell.Exosomes released by specific cell types, such as stem cells, transport bioactive molecules associated with the protection, regeneration, and promotion of blood vessel formation in injured tissues.Further experiments revealed that BMSC-derived exosomes had suppressive activities in inflammation, oxidative stress, and apoptosis, while promoting the proliferation of HG-exposed Müller cells.Additionally, they noted that reintroduction of miR-486-3p increased the beneficial effects of exosome therapy, whereas the upregulation of TLR4 counteracted these  diabetes and associated complications, including DR (Fig. 6).

Conclusion
Diabetes mellitus is a prevalent medical condition that has become more common in recent decades, presenting a significant public health challenge in the twenty-first century.DM serves as a primary contributor to the development of DR, a diabetes complication characterized by the obstruction of ocular blood vessels due to elevated glucose levels, leading to inflammation and leakage of blood or fluids, ultimately resulting in significant ocular damage.The development of DR may involve apoptotic cell death in retinal cells as part of its pathogenesis.Here, our research has shown that key signaling pathways such as PI3K/AKT/mTOR, TLR4/NF-κB, and SIRT1 play significant roles in the apoptosis of retinal cells and the pathogenesis of DR.Notably, an increasing number of research has revealed that ncRNAs can regulate apoptosis in renal cells through various mechanisms.In this context, a recent study has shown that suppressing miR-93 can reduce cell apoptosis and ROS levels in the retinas of DR rats by increasing Nrf2 expression and body's anti-oxidative stress levels (Yin et al. 2019).As well, a recent study has suggested that suppressing miR-93 reduces cell apoptosis and ROS levels in the retinas of DR rats by enhancing the expression of Nrf2 and increasing the body's anti-oxidative stress levels.Furthermore, overexpression of miR-20b-3p, through targeting TXNIP, has been shown to improve visual function and mitigate inflammatory responses, cell apoptosis, vascular permeability, microvascular damage, and angiogenesis in DR rats.These effects contribute to the mitigation of DR progression.These findings suggest miR-20b-3p/ TXNIP axis as a novel therapeutic possibilities for DR management (Wang et al. 2020c).Notably, multiple lncRNAs and circRNAs such as MALAT1, SNHG4, FLG-AS1, SNHG16, circ-0000615, circ-NNT, circFTO, circPAG1, and so forth functions as ceRNAs, targeting microRNAs and influencing the retinal cells apoptosis in DR.Additionally, our findings in the final section reveal that ncRNA-mediated apoptosis modulation via herbal medicine and stem cells-derived exosomes could be exploited to prevent and manage DR (Fig. 7).

Fig. 1 Table 1
Fig. 1 Schematic representation illustrating the pivotal role of apoptosis in the progression of diabetic retinopathy.This visual representation aims to provide a comprehensive overview of

Fig. 2
Fig. 2 Integrative representation elucidating the pivotal signaling pathways orchestrating retinal cell apoptosis in the progression of DR.The diagram highlights key pathways, includ- of each other.While increased LUADT1 xpression correlated with an elevation of peroxiredoxin 3 (PRX3) levels, miR-383 overexpression conversely led to decreased PRX3 expression.Their analysis of cellular apoptosis demonstrated that upregulation of PRX3 and LUADT1 resulted in a decrease in apoptotic cell cell death.Conversely, miR-383 exhibited an opposing effect, mitigating the apoptotic impact of PRX3 and LUADT1 overexpression.Therefore, LUADT1 downregulation observed in DR appears to modulate the PRX3/miR-383 axis, thereby attenuating cell apoptosis and suggesting the LUADT1/miR-383/PRX3 pathway as a potential avenue for therapeutic development in DR (Dai et al. 2022).

Fig. 3
Fig. 3 Illustration depicting the intricate network of apoptosisrelated miRNAs implicated in the pathogenesis of DR.The visual narrative highlights key miRNAs and their regulatory Yu et al. elucidated   the function of the lncRNA SNHG4 and its interaction with miR-200b in DR.Initially, they observed a downregulation of SNHG4 in DR.They provided evidence of a direct interaction between SNHG4 and miR-200b, with the upregulation of miR-200b not exerting any significant impact on the expression of SNHG4 in ARPE-19.In contrast, they observed that SNHG4 upregulation resulted in the increased expression of Oxr1, which is a target of miR-200b.Furthermore, their analysis of cell apoptosis revealed that miR-200b overexpression elevated the rate of apoptosis in ARPE-19 cells subjected to HG treatment.They

Fig. 4
Fig. 4 Comprehensive depiction of the intricate LncRNA/ miRNA/mRNA regulatory axis orchestrating apoptosis in DR.The illustration highlights the dynamic interactions among Consistent with these findings, Li et al. explored the functional significance of exosomal miR-17-3 p in modulating apoptosis in a mice model of DR.They firstly generated a diabetic mouse model.Following Page 25 of 32 53 Vol.: (0123456789) this, exosomes derived from human umbilical cord mesenchymal stem cells (hucMSCs) were isolated and enriched with miR-17-3p before being administered to the mice.A reduction in retinal miR-17-3p expression was observed in DR mice.Conversely, STAT1 expression was increased, suggesting that STAT1 may be a direct target miR-17-3p.Exosome administration enriched with miR-17-3p improved metabolic parameters (reduced blood glucose and HbA1c levels, increased body weight, hemoglobin content, and GS levels) in DR mice.Additionally, it dampened inflammation and vascular dysfunction (decreased inflammatory factors and VEGF levels), mitigated oxidative damage, and suppressed retinal cell apoptosis.These effects were achieved through the inhibition of STAT1 by miR-17-3p delivered via exosomes.In this manner, hucMSCsderived exosomes deliver miR-17-3p to suppress inflammatory responses and apoptosis in DR mice, acting through the downregulation of STAT1 (Li et al. 2021).Furthermore, Li et al. sought to elucidate the mechanisms underlying the impact of BMSCsderived exosomal miR-486-3p in DR.In their study, miR-486-3p exhibited low expression levels, whereas NF-κB and TLR4 exhibited elevated expression levels in HG-exposed Müller cells.They also identified a direct interaction between miR-486-3p and TLR4.The results suggested that increasing miR-486-3p expression or decreasing TLR4 levels resulted in diminished apoptosis, while enhancing the proliferation in Müller cells subjected to HG conditions.

Fig. 5
Fig. 5 Integrated illustration unraveling the intricate Cir-cRNA/miRNA/mRNA regulatory axis governing apoptosis in DR.The visual narrative captures the dynamic interplay

Fig. 6
Fig. 6 Comprehensive depiction of therapeutic strategies targeting apoptosis in DR, spanning from traditional herbal medicines (BAE, AS-4, APS BY) to innovative exosomal miRNA-based interventions.The visual narrative elucidates
(Chen et al. 2017aarget of miR-126, and overexpression of miR-126 decreased IL-17A.In this manner, miR-126 enhances cell growth and prevents cellular apoptosis in HRECs exposed to HG by stimulating the PI3K-−/− ) OIR retinas when compared to wild-type (WT) OIR retinas, thereby confirming the involvement of miR-21 in the downregulation of PPARα in retinal tissue subjected to ischemic conditions.Furthermore, a notable reduction in retinal cell apoptosis, as evidenced by reduced DNA fragmentation, was observed in miR-21 −/− OIR retinas in comparison to WT OIR retinas.These results indicate that the miR-21 silencing mitigated the reduction of PPARα induced by ischemia and improved the condition of retinal apoptosis.In summary, downregulation of PPARα and the induction of apoptosis in DR are, to some extent, mediated by the heightened expression of miR-21 within the diabetic retina(Chen et al. 2017a).